The role of eukaryotic RNA polymerases in site selection on the DNA template will be examined by template binding assays with the RNA polymerase and putative sigma-function proteins. Particularly, the RNA polymerase II will be focused on with respect to template binding by developing an assay to quantitate this enzyme in the presence of multiple other proteins. This assay is based upon the binding of the mushroom toxin, amanitin to the RNA polymerase II. The binding affinity and steric requirements of the amanitin-RNA polymerase complex will be studied in order to design a sensitive and specific assay. This particular assay then will serve to examine the number, distribution, and binding affinity of RNA polymerase II binding sites in high molecular weight DNA and chromatin. The role of putative sigma-functions on RNA polymerase-DNA interactions will be examined by studying the effect of fractionated cell proteins on affinity of RNA polymerase-DNA interaction.